Biochem Exam 2 Lecture 1 PDF

Biochem Exam 2 Lecture 1 -Proteins are organic biomolecules responsible for key biological processes: they are required for replication, maintaining homeostasis, structural support, cell-cell signaling, and developmental coordination among many other functions. -Nucleic acids code for proteins in a...

Biochem Exam 2 Lecture 1 -Proteins are organic biomolecules responsible for key biological processes: they are required for replication, maintaining homeostasis, structural support, cell-cell signaling, and developmental coordination among many other functions. -Nucleic acids code for proteins in a LINEAR fashion: the nucleotide triplet code corresponds linearly to one amino acid after decoding by the ribosome. -The LINEAR sequence of amino acids (AKA polypeptide) is called the PRIMARY STRUCTURE -This linear sequence adopts SECONDARY regular structures (alpha helices and beta sheets) as the protein folds during synthesis. -The secondary structures interact and form the final TERTIARY structure: the final folded form of the complete polypeptide. -Multiple proteins can form larger structures and complexes (e.g. homodimers, heterodimers, tetramers, etc.) this is the QUATERNARY structure. -Well-folded proteins in stable complexes can be crystalized and subjected to X-ray diffraction to determine their 3-dimensional structure to Angstrom (sub-nanometer) resolution. -Protein Structure -> Protein function, and the specific chemistry of the functional R groups of the amino acids can contribute to protein-driven enzymatic catalysis. -“Functional groups”: term from organic chemistry that denotes a group of atoms that have similar chemical properties when they appear in a compound/molecule. E.g. alcohol, aldehyde, ketone. -The main functional groups when thinking about amino acids and proteins are: carboxyl ( OH-C=o), amino (NH2) and the variable R group. -Proteins interact with other proteins, with nucleic acids, lipids and polysaccharides. These interactions help define the biology of the cell. -Most proteins fold into one stable configuration, but this configuration can undergo CONFORMATIONAL CHANGE: a shift in the arrangement of atoms which can alter the protein’s functionality and interactions. There are 20 standard amino acids used by all organisms on Earth. Consider it the alphabet for constructing proteins. This amino acid alphabet is defined by the ‘R group’ on the amino acid, shown here in the red circle, connected to the alpha carbon. - Chirality is handedness. Amino acids have one chiral center, and can be L (Laevorotatory) or D (Dextrorotatory ) isomers. - All life on earth uses L-amino acids: “What is the basis for the preference for L amino acids? The answer has been lost to evolutionary history. It is possible that the preference for L over D amino acids was a consequence of a chance selection.” -Stryer Chirality as a term was first used by Lord Kelvin (William Thomson): “I call any geometrical figure, or group of points, chiral, and say that it has chirality if its image in a plane mirror, ideally realized, cannot be brought to coincide with itself.” Lord Kelvin was not being strictly formal- what he meant by “brought to coincide with itself” was “through rotation and translation cannot be superimposed”. ENANTIOMERS: Affect rotation of polarized light. (You will learn about polarized light and the nature of electromagnetic waves in physics, we will not review it here). Racemic mixture: equal parts of L and R, do not rotate polarized light. Researchers have been able to create entire synthetic systems that use R-amino acids and nucleic acids of opposite chirality. These systems generate biomolecules that are resilient to degradation and have different properties than standard biomolecules. A future goal might be to create entire “mirror image” lifeforms! -In aqueous solution (at terrestrial pressure and temperature ranges which describes the cellular and extracellular conditions relevant to biochemistry) amino acids exist as ZWITTERIONS (from the German word “zwitter” meaning hermaphrodite), which is a chemistry term meaning ‘dipolar ion’: an ion that contains an equal number of positively and negatively charged functional groups (meaning that in aggregate it is electrically neutral, but has non-neutral sides). -Outside of physiological pH range, the non-dipolar forms start to predominate: both sides protonated at low (acidic) pH, and both sides deprotonated at high (basic) pH. -Remember pH tells you the concentration of free H+ (proton) ions in solution! -Highlighted: three letter abbreviations in red are not the first three letters of the amino acid; a different abbreviation was chosen to avoid ambiguity and confusion. -Green boxes: amino acids we’ve into before (SERCA and COX-1) -NOTE Regarding Asx (B) and Glx (Z): These two pairs of amino acids can be ambiguous in peptide sequences because Asp/Asn and Glu/Gln differs only by a terminal amide (-NH2) group in the side chain, and this amide group can be spontaneously lost from proteins by a deamidation reaction. Hence, protein sequences obtained directly from protein frequently contains Asx/Glx entries. -The linear sequence of amino acids is important. The SEQUENCE specifies the STRUCTURE which in turn specifies the FUNCTION. Sequence -> Structure -> Function. st -Insulin was the first protein to be fully sequenced (1 Nobel prize for Sanger). -Many other proteins were then isolated and sequenced via Edman degradation. -Knowledge of protein sequences allowed the Central Dogma (DNA->RNA->Protein) to be established since DNA and RNA triple bases could be assigned to known peptides. -Shown here is the linear sequence for the gene TKTL1 in humans. -Although each of the 20 R groups have distinct properties, you can group them together functionally, (roughly analogously to how the letter “c” and the letter ”k” can serve a similar sound function in the English language). -Most amino acids fall under the category of hydrophobic amino acids, the R group contains alkane chains that are thermodynamically pushed away from water molecules (shells of hydration are low entropy and thus energetically unfavored). -Methionine is often the very first amino acid in a protein due to the start codon of protein synthesis being ATG (Methionine). This is true in both prokaryotes and eukaryotes. -Proline has an unique structural property among all the R groups, which is that it connects back to the amino group (forming a pyrrolidine ring). -NOTICE: Glycine is not chiral!!! -- 5 of the 9 are aliphatic: hydrobophic and uncharged open-chain carbon atoms. Glycine sometimes counted as aliphatic, depending on the source. Polar amino acids; “polar” means a partial charge: the distribution of electrons is uneven over the surface. For example, the –OH group in serine has higher electronegativity than hydrogen or carbon, so there’s more electron density there. This attracts water molecules and makes the association with water thermodynamically favorable. Therefore, many of the polar group can be seen as “hydrophilic” compared to the hydrophobic groups. Two cysteine residues can form covalent bonds called disulfide bridges. -Cysteine can form covalent bonds called disulfide bridges. This can stabilize a conformation of a protein that would normally be unstable or free to rotate, essentially acting as a glue between potentially distant amino acid residues. -Even though cysteine is hydrophilic, it is often not on the surface of proteins due to its reactivity. -Highly hydrophilic -Histidine is often found near the active site of enzymes, due to the property of the imidazole group to readily gain and lose H+ ions at physiological pH (7.3) -Note “aspartate” and “glutamate” are EQUIVALENT to “aspartic acid” and “glumatic acid”. Although the terms are often used interchangeably by biologists, the –ate form refers specifically to the ionic form (shown on this slide). Under physiological conditions (pH 7.4) in proteins the side chain usually occurs as the negatively charged aspartate form. -Glutamate is part of what gives certain foods “umami” flavor (“savoriness”) associated with meatiness. -pKa : chemistry term which means the pH at which 50% of the substance is deprotonated. In all amino acids, the terminal amino and carboxyl groups can be protonated or deprotonated. Since the pKa of a terminal amino group is 8.0, and physiological pH is lower (~7.0, more acidic), we can deduce that most carboxyl groups will be protonated at physiological conditions. This is a dynamic equilibrium state in which any given group may be protonated or deprotonated, but on average at any given time, most will be protonated. -The opposite is true for the carboxyl group, with pKa of 3.1, most of the time it will be deprotonated. When these properties are collected together into a table, it looks like this: Hydrophobic (9) Glycine Alanine (ALIPHATIC) Valine (ALIPHATIC) Leucine (ALIPHATIC) Isoleucine (ALIPHATIC) Methionine (ALIPHATIC) Proline Tryptophan Phenylalanine Polar (6) Serine Threonine Tyrosine (IONIZABLE) Asparagine Glutamine Cysteine (IONIZABLE) Positively Charged (3) Lysine (IONIZABLE) Arginine (IONIZABLE) Histidine (IONIZABLE) Negatively Charged (2) Aspartate (IONIZABLE) Glutamate (IONIZABLE) Remember, if it is ionizable, that R-group has a pkA associated with gaining or losing an H+ ion. st There’s also a “secret” (non-standard) 21 amino acid that is encoded by the UGA stop codon in a small number of proteins (including in humans) called selenocysteine. Earlier we looked at the sequence of human TKTL1. This study provides evidence that a single amino acid change between Neanderthals and modern humans may explain some of our cognitive capacities. The emphasis here is that single amino acid changes in large proteins can have substantial effects on the organism. REMEMBER: These single amino acids changes come from mutations in DNA bases, and are one of the drivers of evolutionary change. -An amino acid is considered “essential” if it cannot be synthesized quickly enough by an organism to meet its biological demand, and must therefore be obtained from diet. This is dependent on the organism in question. Humans do not have to obtain arginine from their diet, but cats do. -Enzymes necessary for constructing a specific amino acid in response to demand may be lost through evolutionary time if there is no selection pressure to maintain them (cats that don’t hunt starve before suffering from arginine deficiency, therefore the presence of dietary arginine is “assumed”). Amino Acids have biological functions other than just being parts of proteins. They are important for maintaining nitrogen homeostasis, they can be metabolized as a source of energy, and can function as signaling molecules and neurotransmitters. -Leucine can act as a direct modulator of the mTORC1 complex, which is a key regulator of cell growth and division. -This is a kind of nutrient sensing: ample amounts of Leucine signal that there is an abundance of resources for growth. It negatively inhibits mTOR which itself negatively inhibits growth and promotes autophagy (cellular recycling for energy conservation). - There is evidence that phenylalanine can also act as an appetite modulator with lactic acid in response to exercise: the intriguing implication is that exercise can actually reduce appetite despite the energy expenditure! Unmodified amino acids can be neurotransmitters (glutamate is the main excitatory neurotransmitter in the mammalian nervous system) or modified into different neurotransmitter (glutamate can be modified into GABA, which is the main inhibitory neurotransmitter). -SOD1 is necessary to control the build-up of damaging free radicals associated with the electron transport chain and oxidative phosphorylation (you will learn about those in future lectures) -Single amino acid substitutions can have severe deleterious consequences. -Here are single amino acid substitutions cause by DNA mutations which can substantially predispose their carriers to ALS/motor neuron disease. -ALS highlights the incredible complexity of linking genetics to disease phenotypes: these mutations vary in penetrance, time to onset of disease, and the exact way the disease progresses. -Red boxes highlight the mutations that have a defined disease progression : e.g. the H46R mutation is “relatively benign” with slower progression. -Different amino acid substitutions may have different effects on the protein’s stability, localization, interactions, and enzymatic function.